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1.
Am J Med Genet A ; 191(12): 2837-2842, 2023 12.
Artigo em Inglês | MEDLINE | ID: mdl-37554039

RESUMO

Left main coronary artery ostial atresia (LMCAOA) is an extremely rare condition. Here, we report the case of a 14-year-old boy with Noonan syndrome-like disorder in whom LMCAOA was detected following cardiopulmonary arrest. The patient had been diagnosed with Noonan syndrome-like disorder with a pathogenic splice site variant of CBL c.1228-2 A > G. He suddenly collapsed when he was running. After administering two electric shocks using an automated external defibrillator, the patient's heartbeat resumed. Cardiac catheterization confirmed the diagnosis of LMCAOA. Left main coronary artery angioplasty was performed. The patient was discharged without neurological sequelae. Brain magnetic resonance imaging revealed asymptomatic Moyamoya disease. In addition, RNF213 c.14429 G > A p.R4810K was identified. There are no reports on congenital coronary malformations of compound variations of RNF213 and CBL. In contrast, the RNF213 p.R4810K polymorphism has been established as a risk factor for angina pectoris and myocardial infarction in adults, and several congenital coronary malformations due to genetic abnormalities within the RAS/MAPK signaling pathway have been reported. This report aims to highlight the risk of sudden death in patients with RASopathy and RNF213 p.R4810K polymorphism and emphasize the significance of actively searching for coronary artery morphological abnormalities in these patients.


Assuntos
Anormalidades Múltiplas , Parada Cardíaca , Doença de Moyamoya , Síndrome de Noonan , Adulto , Masculino , Humanos , Criança , Adolescente , Vasos Coronários/diagnóstico por imagem , Vasos Coronários/metabolismo , Síndrome de Noonan/complicações , Síndrome de Noonan/diagnóstico , Síndrome de Noonan/genética , Predisposição Genética para Doença , Adenosina Trifosfatases/genética , Ubiquitina-Proteína Ligases/genética , Doença de Moyamoya/genética , Parada Cardíaca/genética
2.
Circ Arrhythm Electrophysiol ; 16(3): e011387, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36866681

RESUMO

BACKGROUND: CaM (calmodulin) is a ubiquitously expressed, multifunctional Ca2+ sensor protein that regulates numerous proteins. Recently, CaM missense variants have been identified in patients with malignant inherited arrhythmias, such as long QT syndrome and catecholaminergic polymorphic ventricular tachycardia (CPVT). However, the exact mechanism of CaM-related CPVT in human cardiomyocytes remains unclear. In this study, we sought to investigate the arrhythmogenic mechanism of CPVT caused by a novel variant using human induced pluripotent stem cell (iPSC) models and biochemical assays. METHODS: We generated iPSCs from a patient with CPVT bearing CALM2 p.E46K. As comparisons, we used 2 control lines including an isogenic line, and another iPSC line from a patient with long QT syndrome bearing CALM2 p.N98S (also reported in CPVT). Electrophysiological properties were investigated using iPSC-cardiomyocytes. We further examined the RyR2 (ryanodine receptor 2) and Ca2+ affinities of CaM using recombinant proteins. RESULTS: We identified a novel de novo heterozygous variant, CALM2 p.E46K, in 2 unrelated patients with CPVT accompanied by neurodevelopmental disorders. The E46K-cardiomyocytes exhibited more frequent abnormal electrical excitations and Ca2+ waves than the other lines in association with increased Ca2+ leakage from the sarcoplasmic reticulum via RyR2. Furthermore, the [3H]ryanodine binding assay revealed that E46K-CaM facilitated RyR2 function especially by activating at low [Ca2+] levels. The real-time CaM-RyR2 binding analysis demonstrated that E46K-CaM had a 10-fold increased RyR2 binding affinity compared with wild-type CaM which may account for the dominant effect of the mutant CaM. Additionally, the E46K-CaM did not affect CaM-Ca2+ binding or L-type calcium channel function. Finally, antiarrhythmic agents, nadolol and flecainide, suppressed abnormal Ca2+ waves in E46K-cardiomyocytes. CONCLUSIONS: We, for the first time, established a CaM-related CPVT iPSC-CM model which recapitulated severe arrhythmogenic features resulting from E46K-CaM dominantly binding and facilitating RyR2. In addition, the findings in iPSC-based drug testing will contribute to precision medicine.


Assuntos
Células-Tronco Pluripotentes Induzidas , Síndrome do QT Longo , Taquicardia Ventricular , Humanos , Calmodulina/genética , Calmodulina/metabolismo , Miócitos Cardíacos/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Taquicardia Ventricular/metabolismo , Arritmias Cardíacas , Síndrome do QT Longo/genética , Síndrome do QT Longo/metabolismo , Cálcio/metabolismo , Mutação
3.
Med Mol Morphol ; 54(3): 259-264, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33651170

RESUMO

A 16-year-old Japanese man was admitted to our hospital because of syncope during exercise. His father and his younger brother had permanent pacemaker implantation because of sick sinus syndrome. Several examinations revealed first-degree atrioventricular block, complete right bundle branch block, sick sinus syndrome, and ventricular tachycardia with normal cardiac function. As no abnormalities were evident on coronary angiography, right ventricular endomyocardial biopsy was performed. It showed myocardial disarrangement and lipofuscin accumulation in hypertrophic myocytes. Moreover, electron microscopy showed a few degenerative myocytes, Z-band streaming, disarrangement, increased small capillaries with Weibel-Palade bodies in endothelial cells, and endothelial proliferations. Genetic analysis of the proband, his father, and his younger brother revealed a missense mutation, D1275N, in SCN5A, a gene which encodes sodium ion channel protein, are related to cardiomyopathy and arrhythmia. The proband was diagnosed with a cardiac conduction defect (CCD) and underwent permanent pacemaker implantation. These pathological findings suggest various myocardial changes presented in CCD patients with a missense mutation, D1275N, in SCN5A.


Assuntos
Doença do Sistema de Condução Cardíaco/genética , Mutação de Sentido Incorreto , Miocárdio/patologia , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Adolescente , Arritmias Cardíacas/genética , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/patologia , Doença do Sistema de Condução Cardíaco/metabolismo , Doença do Sistema de Condução Cardíaco/patologia , Doença do Sistema de Condução Cardíaco/terapia , Humanos , Masculino , Linhagem
4.
Circ J ; 82(11): 2707-2714, 2018 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-30078822

RESUMO

BACKGROUND: Mutation in the lamin A/C gene (LMNA) is associated with several cardiac phenotypes, such as cardiac conduction disorders (CCD), atrial arrhythmia (AA), malignant ventricular arrhythmia (MVA) and left ventricular dysfunction (LVD), leading to sudden cardiac death (SCD) and/or end-stage heart failure. We investigated how these phenotypes are associated with each other and which of them are most important for total mortality. Methods and Results: A multicenter registry included 110 LMNA mutation carriers (age, 43±15 years, male: 62%) from 60 families. After genetic diagnosis of LMNA mutation (missense: 27%, non-missense: 73%), patients or subjects were followed to evaluate the manifestations of their phenotypes and the risk of total mortality; 90 patients could be followed (median: 5 [0-35] years). Prevalence of the 4 clinical phenotypes was significantly increased during follow-up. Among these phenotypes, AA was significantly associated with MVA. CCD was significantly associated with LVD. LVD, meanwhile, was significantly associated with CCD and MVA. Male sex was significantly associated with MVA. Furthermore, during follow-up, 17 patients died: 12 end-stage heart failure, 4 SCD and 1 stroke. LVD was the only independent predictor for all-cause death (OR: 41.7, 95% CI: 4.1-422.3; P=0.0016). CONCLUSIONS: Several cardiac phenotypes were age-dependently increased in LMNA mutation carriers, suggesting that ICD or CRT-D could suppress SCD after middle age; however, LVD leading to end-stage heart failure was the only independent predictor for total mortality.


Assuntos
Cardiopatias/genética , Cardiopatias/mortalidade , Lamina Tipo A/genética , Mutação , Sistema de Registros , Adulto , Fatores Etários , Feminino , Seguimentos , Humanos , Japão/epidemiologia , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores Sexuais
5.
Circ Cardiovasc Genet ; 10(6)2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-29237675

RESUMO

BACKGROUND: Mutations in LMNA (lamin A/C), which encodes lamin A and C, typically cause age-dependent cardiac phenotypes, including dilated cardiomyopathy, cardiac conduction disturbance, atrial fibrillation, and malignant ventricular arrhythmias. Although the type of LMNA mutations have been reported to be associated with susceptibility to malignant ventricular arrhythmias, the gene-based risk stratification for cardiac complications remains unexplored. METHODS AND RESULTS: The multicenter cohort included 77 LMNA mutation carriers from 45 families; cardiac disorders were retrospectively analyzed. The mean age of patients when they underwent genetic testing was 45±17, and they were followed for a median 49 months. Of the 77 carriers, 71 (92%) were phenotypically affected and showed cardiac conduction disturbance (81%), low left ventricular ejection fraction (<50%; 45%), atrial arrhythmias (58%), and malignant ventricular arrhythmias (26%). During the follow-up period, 9 (12%) died, either from end-stage heart failure (n=7) or suddenly (n=2). Genetic analysis showed truncation mutations in 58 patients from 31 families and missense mutations in 19 patients from 14 families. The onset of cardiac disorders indicated that subjects with truncation mutations had an earlier occurrence of cardiac conduction disturbance and low left ventricular ejection fraction, than those with missense mutations. In addition, the truncation mutation was found to be a risk factor for the early onset of cardiac conduction disturbance and the occurrence of atrial arrhythmias and low left ventricular ejection fraction, as estimated using multivariable analyses. CONCLUSIONS: The truncation mutations were associated with manifestation of cardiac phenotypes in LMNA-related cardiomyopathy, suggesting that genetic analysis might be useful for diagnosis and risk stratification.


Assuntos
Cardiomiopatias/genética , Predisposição Genética para Doença/genética , Lamina Tipo A/genética , Mutação , Adulto , Idoso , Arritmias Cardíacas/genética , Arritmias Cardíacas/fisiopatologia , Cardiomiopatias/mortalidade , Cardiomiopatias/fisiopatologia , Saúde da Família , Feminino , Sistema de Condução Cardíaco/fisiopatologia , Heterozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Fenótipo , Estudos Retrospectivos , Fatores de Risco , Taxa de Sobrevida
6.
J Am Coll Cardiol ; 70(3): 358-370, 2017 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-28705318

RESUMO

BACKGROUND: Inherited cardiac conduction disease is a rare bradyarrhythmia associated with mutations in various genes that affect action potential propagation. It is often characterized by isolated conduction disturbance of the His-Purkinje system, but it is rarely described as a syndromic form. OBJECTIVES: The authors sought to identify the genetic defect in families with a novel bradyarrhythmia syndrome associated with bone malformation. METHODS: The authors genetically screened 15 European cases with genotype-negative de novo atrioventricular (AV) block and their parents by trio whole-exome sequencing, plus 31 Japanese cases with genotype-negative familial AV block or sick sinus syndrome by targeted exon sequencing of 457 susceptibility genes. Functional consequences of the mutation were evaluated using an in vitro cell expression system and in vivo knockout mice. RESULTS: The authors identified a connexin-45 (Cx45) mutation (p.R75H) in 2 unrelated families (a de novo French case and a 3-generation Japanese family) who presented with progressive AV block, which resulted in atrial standstill without ventricular conduction abnormalities. Affected individuals shared a common extracardiac phenotype: a brachyfacial pattern, finger deformity, and dental dysplasia. Mutant Cx45 expressed in Neuro-2a cells showed normal hemichannel assembly and plaque formation. However, Lucifer yellow dye transfer and gap junction conductance between cell pairs were severely impaired, which suggested that mutant Cx45 impedes gap junction communication in a dominant-negative manner. Tamoxifen-induced, cardiac-specific Cx45 knockout mice showed sinus node dysfunction and atrial arrhythmia, recapitulating the intra-atrial disturbance. CONCLUSIONS: Altogether, the authors showed that Cx45 mutant p.R75H is responsible for a novel disease entity of progressive atrial conduction system defects associated with craniofacial and dentodigital malformation.


Assuntos
Bloqueio Atrioventricular/etiologia , Conexinas/genética , DNA/genética , Deformidades Dentofaciais/complicações , Mutação , Adolescente , Adulto , Animais , Bloqueio Atrioventricular/genética , Bloqueio Atrioventricular/fisiopatologia , Criança , Pré-Escolar , Conexinas/metabolismo , Análise Mutacional de DNA , Deformidades Dentofaciais/genética , Deformidades Dentofaciais/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Eletrocardiografia , Feminino , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Linhagem , Fenótipo , Adulto Jovem
7.
Circ Arrhythm Electrophysiol ; 8(5): 1095-104, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26129877

RESUMO

BACKGROUND: Few rare variants in atrial fibrillation (AF)-associated genes have been functionally characterized to identify a causal relationship between these variants and development of AF. We here sought to determine the clinical effect of rare variants in AF-associated genes in patients with lone AF and characterized these variants electrophysiologically and bioinformatically. METHODS AND RESULTS: We screened all coding regions in 12 AF-associated genes in 90 patients with lone AF, with an onset of 47±11 years (66 men; mean age, 56±13 years) by high-resolution melting curve analysis and DNA sequencing. The potassium and sodium currents were analyzed using whole-cell patch clamping. In addition to using 4 individual in silico prediction tools, we extended those predictions to an integrated tool (Combined Annotation Dependent Depletion). We identified 7 rare variants in KCNA5, KCNQ1, KCNH2, SCN5A, and SCN1B genes in 8 patients: 2 of 8 probands had a family history of AF. Electrophysiological studies revealed that 2 variants showed a loss-of-function, and 4 variants showed a gain-of-function. Five of 6 variants with electrophysiological abnormalities were predicted as pathogenic by Combined Annotation Dependent Depletion scores. CONCLUSIONS: In our cohort of patients with lone AF, 7 rare variants in cardiac ion channels were identified in 8 probands. A combination of electrophysiological studies and in silico predictions showed that these variants could contribute to the development of lone AF, although further in vivo study is necessary to confirm these results. More than half of AF-associated rare variants showed gain-of-function behavior, which may be targeted using genotype-specific pharmacological therapy.


Assuntos
Fibrilação Atrial/genética , Variação Genética , Fibrilação Atrial/fisiopatologia , Canal de Potássio ERG1 , Técnicas Eletrofisiológicas Cardíacas , Canais de Potássio Éter-A-Go-Go/genética , Feminino , Predisposição Genética para Doença , Genótipo , Humanos , Canais Iônicos/genética , Canal de Potássio KCNQ1/genética , Canal de Potássio Kv1.5/genética , Masculino , Pessoa de Meia-Idade , Mutação/genética , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Técnicas de Patch-Clamp , Subunidade beta-1 do Canal de Sódio Disparado por Voltagem/genética
8.
Heart Vessels ; 29(3): 422-6, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-23836069

RESUMO

We present a case of a 9-month-old girl in whom malignant polymorphic ventricular tachycardia (VT) was successfully controlled by radiofrequency catheter ablation under guidance with a three-dimensional mapping system. The VTs originated from the left ventricular lateral wall, left ventricular anterior wall, and left ventricular apex. At least six types of VTs were documented during the electrophysiology study. All VTs were successfully controlled after two sessions of radiofrequency catheter ablation, and she was discharged from our hospital on propranolol, mexiletine, flecainide, and aprindine.


Assuntos
Ablação por Cateter , Frequência Cardíaca , Taquicardia Ventricular/cirurgia , Antiarrítmicos/uso terapêutico , Eletrocardiografia , Técnicas Eletrofisiológicas Cardíacas , Feminino , Humanos , Lactente , Taquicardia Ventricular/diagnóstico , Taquicardia Ventricular/fisiopatologia , Resultado do Tratamento
9.
PLoS Genet ; 9(4): e1003364, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23593010

RESUMO

Unexplained cardiac arrest (UCA) with documented ventricular fibrillation (VF) is a major cause of sudden cardiac death. Abnormal sympathetic innervations have been shown to be a trigger of ventricular fibrillation. Further, adequate expression of SEMA3A was reported to be critical for normal patterning of cardiac sympathetic innervation. We investigated the relevance of the semaphorin 3A (SEMA3A) gene located at chromosome 5 in the etiology of UCA. Eighty-three Japanese patients diagnosed with UCA and 2,958 healthy controls from two different geographic regions in Japan were enrolled. A nonsynonymous polymorphism (I334V, rs138694505A>G) in exon 10 of the SEMA3A gene identified through resequencing was significantly associated with UCA (combined P = 0.0004, OR 3.08, 95%CI 1.67-5.7). Overall, 15.7% of UCA patients carried the risk genotype G, whereas only 5.6% did in controls. In patients with SEMA3A(I334V), VF predominantly occurred at rest during the night. They showed sinus bradycardia, and their RR intervals on the 12-lead electrocardiography tended to be longer than those in patients without SEMA3A(I334V) (1031±111 ms versus 932±182 ms, P = 0.039). Immunofluorescence staining of cardiac biopsy specimens revealed that sympathetic nerves, which are absent in the subendocardial layer in normal hearts, extended to the subendocardial layer only in patients with SEMA3A(I334V). Functional analyses revealed that the axon-repelling and axon-collapsing activities of mutant SEMA3A(I334V) genes were significantly weaker than those of wild-type SEMA3A genes. A high incidence of SEMA3A(I334V) in UCA patients and inappropriate innervation patterning in their hearts implicate involvement of the SEMA3A gene in the pathogenesis of UCA.


Assuntos
Parada Cardíaca , Coração , Semaforina-3A/genética , Fibrilação Ventricular , Adulto , Idoso , Feminino , Coração/inervação , Coração/fisiopatologia , Parada Cardíaca/genética , Parada Cardíaca/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Miocárdio/metabolismo , Fatores de Risco , Semaforina-3A/metabolismo , Sistema Nervoso Simpático/metabolismo , Sistema Nervoso Simpático/fisiopatologia , Fibrilação Ventricular/genética , Fibrilação Ventricular/metabolismo , Fibrilação Ventricular/fisiopatologia
10.
Cardiovasc Res ; 95(4): 419-29, 2012 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-22739119

RESUMO

AIMS: Long QT syndrome (LQTS) is an inheritable and life-threatening disease; however, it is often difficult to determine disease characteristics in sporadic cases with novel mutations, and more precise analysis is necessary for the successful development of evidence-based clinical therapies. This study thus sought to better characterize ion channel cardiac disorders using induced pluripotent stem cells (iPSCs). METHODS AND RESULTS: We reprogrammed somatic cells from a patient with sporadic LQTS and from controls, and differentiated them into cardiomyocytes through embryoid body (EB) formation. Electrophysiological analysis of the LQTS-iPSC-derived EBs using a multi-electrode array (MEA) system revealed a markedly prolonged field potential duration (FPD). The IKr blocker E4031 significantly prolonged FPD in control- and LQTS-iPSC-derived EBs and induced frequent severe arrhythmia only in LQTS-iPSC-derived EBs. The IKs blocker chromanol 293B did not prolong FPD in the LQTS-iPSC-derived EBs, but significantly prolonged FPD in the control EBs, suggesting the involvement of IKs disturbance in the patient. Patch-clamp analysis and immunostaining confirmed a dominant-negative role for 1893delC in IKs channels due to a trafficking deficiency in iPSC-derived cardiomyocytes and human embryonic kidney (HEK) cells. CONCLUSIONS: This study demonstrated that iPSCs could be useful to characterize LQTS disease as well as drug responses in the LQTS patient with a novel mutation. Such analyses may in turn lead to future progress in personalized medicine.


Assuntos
Células-Tronco Pluripotentes Induzidas/metabolismo , Miócitos Cardíacos/metabolismo , Síndrome de Romano-Ward/metabolismo , Potenciais de Ação , Adolescente , Animais , Diferenciação Celular , Reprogramação Celular , Técnicas de Cocultura , Corpos Embrioides/metabolismo , Corpos Embrioides/patologia , Imunofluorescência , Regulação da Expressão Gênica no Desenvolvimento , Predisposição Genética para Doença , Células HEK293 , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/patologia , Células-Tronco Pluripotentes Induzidas/transplante , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/antagonistas & inibidores , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/genética , Canais de Potássio Ativados por Cálcio de Condutância Intermediária/metabolismo , Canal de Potássio KCNQ1/antagonistas & inibidores , Canal de Potássio KCNQ1/genética , Canal de Potássio KCNQ1/metabolismo , Masculino , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Miócitos Cardíacos/transplante , Técnicas de Patch-Clamp , Fenótipo , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores , Canais de Potássio de Abertura Dependente da Tensão da Membrana/genética , Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Síndrome de Romano-Ward/diagnóstico , Síndrome de Romano-Ward/genética , Síndrome de Romano-Ward/patologia , Teratoma/metabolismo , Teratoma/patologia , Fatores de Tempo , Transfecção
11.
Curr Opin Cardiol ; 27(3): 236-41, 2012 May.
Artigo em Inglês | MEDLINE | ID: mdl-22382502

RESUMO

PURPOSE OF REVIEW: Connexins are the pore forming subunits of gap junction channels. They are essential for cardiac action potential propagation. Connexins are modified at the transcriptional or posttranslational levels under pathological states such as cardiac hypertrophy or ischemia, thus contributing to the arrhythmogenic substrate. However, the relation between nucleotide substitutions in the connexin gene and the occurrence of cardiac arrhythmias remains largely unexplored. RECENT FINDINGS: Recent studies have reported an association between nucleotide substitutions in the connexin40 (Cx40) and connexin43 (Cx43) genes (GJA5 and GJA1, respectively) and cardiac arrhythmias. Of note, however, germline mutations in Cx43 are considered causative of oculodentodigital dysplasia, a pleiotropic syndrome wherein cardiac manifestations are notoriously absent. SUMMARY: Here, we review some of the current knowledge on the association between cardiac connexins and inherited arrhythmias.


Assuntos
Arritmias Cardíacas/genética , Conexina 43/genética , Conexinas/genética , Miocárdio , Potenciais de Ação , Arritmias Cardíacas/etiologia , Arritmias Cardíacas/patologia , Fibrilação Atrial/genética , Cardiomiopatias/genética , Predisposição Genética para Doença , Bloqueio Cardíaco , Humanos , Mutação , Proteína alfa-5 de Junções Comunicantes
12.
Circ Arrhythm Electrophysiol ; 5(1): 163-72, 2012 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-22247482

RESUMO

BACKGROUND: Progressive familial heart block type I (PFHBI) is a hereditary arrhythmia characterized by progressive conduction disturbances in the His-Purkinje system. PFHBI has been linked to genes such as SCN5A that influence cardiac excitability but not to genes that influence cell-to-cell communication. Our goal was to explore whether nucleotide substitutions in genes coding for connexin proteins would associate with clinical cases of PFHBI and if so, to establish a genotype-cell phenotype correlation for that mutation. METHODS AND RESULTS: We screened 156 probands with PFHBI. In addition to 12 sodium channel mutations, we found a germ line GJA5 (connexin40 [Cx40]) mutation (Q58L) in 1 family. Heterologous expression of Cx40-Q58L in connexin-deficient neuroblastoma cells resulted in marked reduction of junctional conductance (Cx40-wild type [WT], 22.2±1.7 nS, n=14; Cx40-Q58L, 0.56±0.34 nS, n=14; P<0.001) and diffuse localization of immunoreactive proteins in the vicinity of the plasma membrane without formation of gap junctions. Heteromeric cotransfection of Cx40-WT and Cx40-Q58L resulted in homogenous distribution of proteins in the plasma membrane rather than in membrane plaques in ≈50% of cells; well-defined gap junctions were observed in other cells. Junctional conductance values correlated with the distribution of gap junction plaques. CONCLUSIONS: Mutation Cx40-Q58L impairs gap junction formation at cell-cell interfaces. This is the first demonstration of a germ line mutation in a connexin gene that associates with inherited ventricular arrhythmias and emphasizes the importance of Cx40 in normal propagation in the specialized conduction system.


Assuntos
Fascículo Atrioventricular/metabolismo , Conexinas/genética , DNA/genética , Bloqueio Cardíaco/genética , Mutação , Biomarcadores/metabolismo , Western Blotting , Fascículo Atrioventricular/fisiopatologia , Doença do Sistema de Condução Cardíaco , Criança , Conexinas/metabolismo , Eletrocardiografia , Técnicas Eletrofisiológicas Cardíacas , Feminino , Predisposição Genética para Doença , Bloqueio Cardíaco/metabolismo , Bloqueio Cardíaco/fisiopatologia , Frequência Cardíaca , Humanos , Imuno-Histoquímica , Masculino , Linhagem , Reação em Cadeia da Polimerase , Prognóstico , Proteína alfa-5 de Junções Comunicantes
13.
Life Sci ; 84(11-12): 380-7, 2009 Mar 13.
Artigo em Inglês | MEDLINE | ID: mdl-19167409

RESUMO

AIMS: In the type 3 long QT syndrome (LQT3), shortening of the QT interval by overdrive pacing is used to prevent life-threatening arrhythmias. However, it is unclear whether accelerated heart rate induced by beta-adrenergic agents produces similar effects on the late sodium current (I(Na)) to those by overdrive pacing therapy. We analyzed the beta-adrenergic-like effects of protein kinase A and fluoride on I(Na) in R1623Q mutant channels. MAIN METHODS: cDNA encoding either wild-type (WT) or R1623Q mutant of hNa(v)1.5 was stably transfected into HEK293 cells. I(Na) was recorded using a whole-cell patch-clamp technique at 23 degrees C. KEY FINDINGS: In R1623Q channels, 2 mM pCPT-AMP and 120 mM fluoride significantly delayed macroscopic current decay and increased relative amplitude of the late I(Na) in a time-dependent manner. Modulations of peak I(Na) gating kinetics (activation, inactivation, recovery from inactivation) by fluoride were similar in WT and R1623Q channels. The effects of fluoride were almost completely abolished by concomitant dialysis with a protein kinase inhibitor. We also compared the effect of pacing with that of beta-adrenergic stimulation by analyzing the frequency-dependence of the late I(Na). Fluoride augmented frequency-dependent reduction of the late I(Na), which was due to preferential delay of recovery of late I(Na). However, the increase in late I(Na) by fluoride at steady-state was more potent than the frequency-dependent reduction of late I(Na). SIGNIFICANCE: Different basic mechanisms participate in the QT interval shortening by pacing and beta-adrenergic stimulation in the LQT3.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Ativação do Canal Iônico , Síndrome do QT Longo/metabolismo , Proteínas Musculares/genética , Canais de Sódio/genética , Potenciais de Ação/efeitos dos fármacos , Linhagem Celular , Clonagem Molecular , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacologia , Proteínas Quinases Dependentes de AMP Cíclico/antagonistas & inibidores , Fluoretos/farmacologia , Humanos , Ativação do Canal Iônico/efeitos dos fármacos , Síndrome do QT Longo/enzimologia , Síndrome do QT Longo/genética , Mutação , Canal de Sódio Disparado por Voltagem NAV1.5 , Técnicas de Patch-Clamp , Inibidores de Proteínas Quinases/farmacologia , Tionucleotídeos/farmacologia , Transfecção
14.
Pediatr Res ; 64(5): 482-7, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18596570

RESUMO

Sudden infant death syndrome (SIDS) is multifactorial and may result from the interaction of a number of environmental, genetic, and developmental factors. We studied three major genes causing long QT syndrome in 42 Japanese SIDS victims and found five mutations, KCNQ1-K598R, KCNH2-T895M, SCN5A-F532C, SCN5A-G1084S, and SCN5A-F1705S, in four cases; one case had both KCNH2-T895M and SCN5A-G1084S. All mutations were novel except for SCN5A-F532C, which was previously detected in an arrhythmic patient. Heterologous expression study revealed significant changes in channel properties of KCNH2-T895M, SCN5A-G1084S, and SCN5A-F1705S, but did not in KCNQ1-K598R and SCN5A-F532C. Our data suggests that nearly 10% of SIDS victims in Japan have mutations of the cardiac ion channel genes similar to in other countries.


Assuntos
Canais de Potássio Éter-A-Go-Go/genética , Canal de Potássio KCNQ1/genética , Proteínas Musculares/genética , Mutação , Miocárdio/metabolismo , Canais de Sódio/genética , Morte Súbita do Lactente/genética , Animais , Povo Asiático/genética , Linhagem Celular , Estudos de Coortes , Canal de Potássio ERG1 , Canais de Potássio Éter-A-Go-Go/metabolismo , Feminino , Predisposição Genética para Doença , Humanos , Lactente , Recém-Nascido , Japão , Canal de Potássio KCNQ1/metabolismo , Cinética , Masculino , Potenciais da Membrana , Proteínas Musculares/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.5 , Canais de Sódio/metabolismo , Morte Súbita do Lactente/etnologia , Transfecção , Xenopus laevis
15.
Heart Rhythm ; 4(4): 516-9, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17399644

RESUMO

BACKGROUND: An association between Brugada syndrome and neurally mediated syncope has been described. Although mutations in SCN5A have been identified in Brugada syndrome, the genetic link between Brugada syndrome and neurally mediated syncope has not been determined. OBJECTIVES: The purpose of the study was to clinically and genetically characterize a man with recurrent syncope that originally was diagnosed as neurally mediated syncope at age 8 years but subsequently manifested as Brugada syndrome at age 17 years. METHODS: The proband underwent clinical examination, which included head-up tilt test, sodium channel provocation test, and electrophysiologic study. Genetic screening of SCN5A was performed for the proband and his family members. The biophysical properties of a mutant SCN5A channel in a heterologous expression system were studied using whole-cell, patch clamp technique. RESULTS: The proband showed positive head-up tilt test, coved-type ST elevation recorded from the third intercostal space, and positive pilsicainide provocation test. Ventricular fibrillation was inducible at programmed electrical stimulation, consistent with characteristics of both Brugada syndrome and neurally mediated syncope. A novel nonsense SCN5A mutation (Q55X) was identified in the proband, his mother, and his asymptomatic brother. The heterologously expressed mutant channel was nonfunctional. CONCLUSION: We genetically determined an SCN5A mutation in a patient showing the combined phenotype of neurally mediated syncope and Brugada syndrome. Neurally mediated syncope and Brugada syndrome may share, at least in part, a common pathophysiologic mechanism.


Assuntos
Síndrome de Brugada/complicações , Síndrome de Brugada/genética , Proteínas Musculares/genética , Canais de Sódio/genética , Síncope Vasovagal/etiologia , Potenciais de Ação , Adolescente , Fatores Etários , Nó Atrioventricular/fisiopatologia , Síndrome de Brugada/fisiopatologia , Códon sem Sentido/genética , Técnicas Eletrofisiológicas Cardíacas , Éxons/genética , Humanos , Masculino , Canal de Sódio Disparado por Voltagem NAV1.5 , Linhagem , Nó Sinoatrial/fisiopatologia
16.
Eur J Neurosci ; 22(4): 793-801, 2005 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16115203

RESUMO

Both tetrodotoxin-sensitive (TTX-S) and TTX-resistant (TTX-R) voltage-dependent Na+ channels are expressed in the human neuroblastoma cell line NB-1, but a gene encoding the TTX-R Na+ channel has not been identified. In this study, we have cloned cDNA encoding the alpha subunit of the TTX-R Na+ channel in NB-1 cells and designated it hNbR1. The longest open reading frame of hNbR1 (accession no. AB158469) encodes 2016 amino acid residues. Sequence analysis has indicated that hNbR1 is highly homologous with human cardiac Nav1.5/SCN5A with > 99% amino acid identity. The presence of a cysteine residue (Cys373) in the pore-loop region of domain I is consistent with the supposition that hNbR1 is resistant to TTX. Analysis of the genomic sequence of SCN5A revealed a new exon encoding S3 and S4 of domain I (exon 6A). In addition, an alternative splicing variant, lacking exon 18, that encodes 54 amino acids in the intracellular loop between domains II and III was found (hNbR1-2; accession no. AB158470). Na+ currents in human embryonic kidney cells (HEK293) transfected with hNbR1 or hNbR1-2 showed electrophysiological properties similar to those for TTX-R I(Na) in NB-1 cells. The IC50 for the TTX block was approximately 8 microM in both variants. These results suggest that SCN5A has a newly identified exon for alternative splicing and is more widely expressed than previously thought.


Assuntos
Variação Genética , Proteínas Musculares/genética , Proteínas/genética , Canais de Sódio/genética , Tetrodotoxina/farmacologia , Sequência de Aminoácidos , Northern Blotting/métodos , Linhagem Celular Tumoral , Cromossomos Humanos Par 3 , Clonagem Molecular/métodos , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Estimulação Elétrica , Éxons , Humanos , Concentração Inibidora 50 , Peptídeos e Proteínas de Sinalização Intracelular , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Potenciais da Membrana/efeitos da radiação , Dados de Sequência Molecular , Proteínas Musculares/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.5 , Neuroblastoma , Técnicas de Patch-Clamp/métodos , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Bloqueadores dos Canais de Sódio/farmacologia , Canais de Sódio/metabolismo , Transfecção/métodos
17.
Circ J ; 66(9): 846-50, 2002 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-12224824

RESUMO

The present study investigated the protective effects of L-cysteine on the oxidation-induced blockade of Na+ channel a-subunits, hH1 (cardiac) and hSkM1 (skeletal), expressed in COS7 cells. Na+ currents were recorded by the whole-cell patch clamp technique (n = 3-7). L-cysteine alone blocked hH1 and hSkM1 in a dose-dependent manner, with saturating L-cysteine block at 3,000 micromol/L. Hg2+, a potent sulfhydryl oxidizing agent, blocked hH1 with a time to 50% inhibition (Time50%) of 20s. Preperfusion of COS7 cells with 100 micromol/L L-cysteine significantly slowed the Hg2+ block of hH1 (Time50% = 179 s). L-cysteine did not prevent Hg2+ block of hSkM1 (Time50% = 37s) or the C373Y hH1 mutant (Time50% = 43s). As for other sulfo-amino acids, homocysteine prevented the Hg2+ block of hH1, with the Time50% (70s) being significantly smaller than that of L-cysteine, whereas methionine did not prevent the Hg2+ block of hH1. L-cysteine did not prevent the Cd2+ block of hH1. These results indicate that L-cysteine selectively acts on heart-specific Cys373 in the P-loop region of hH1 to prevent Cys373 from the oxidation-induced sulfur-Hg-sulfur bridge formation.


Assuntos
Cisteína/farmacologia , Canais de Sódio/efeitos dos fármacos , Relação Dose-Resposta a Droga , Homocisteína/farmacologia , Humanos , Mercúrio/farmacologia , Metionina/farmacologia , Miocárdio/citologia , Oxirredução , Técnicas de Patch-Clamp , Canais de Sódio/genética
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